GSM7118703: CD90posCD55pos, biol rep2; Mus musculus; RNA-Seq
Bones were isolated from limbs and minced with scissors or tweezers. The fragmented bones were incubated in DMEM containing Collagenase P, Liberase TH and DNase I at 37ºC with gentle shaking for 1h. Dissociated cells were collected every 20 min and strained with a 100 μm nylon mesh. Cells were collected by centrifuge and resuspended in FACS buffer (2% FBS/2 mM EDTA/PBS). The isolated cells were stained with antibodies against CD45, Ter119 and CD31. Fixable viability dye (eBioscience) was used to stain dead cells. Cells were sorted directly into SMART-Seq reaction buffer for library construction. RNA libraries for RNA-seq were prepared using SMART-Seq HT Plus Kit (Takara Bio) following manufacturer's protocols.